Haile D J, Rouault T A, Harford J B, Kennedy M C, Blondin G A, Beinert H, Klausner R D
Cell Biology and Metabolism Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892.
Proc Natl Acad Sci U S A. 1992 Dec 15;89(24):11735-9. doi: 10.1073/pnas.89.24.11735.
The translation of ferritin mRNA and degradation of transferrin receptor mRNA are regulated by the interaction of an RNA-binding protein, the iron-responsive element binding protein (IRE-BP), with RNA stem-loop structures known as iron-responsive elements (IREs) contained within these transcripts. IRE-BP produced in iron-replete cells has aconitase (EC 4.2.1.3) activity. The protein shows extensive sequence homology with mitochondrial aconitase, and sequences of peptides prepared from cytosolic aconitase are identical with peptides of IRE-BP. As an active aconitase, IRE-BP is expected to have an Fe-S cluster, in analogy to other aconitases. This Fe-S cluster has been implicated as the region of the protein that senses intracellular iron levels and accordingly modifies the ability of the IRE-BP to interact with IREs. Expression of the IRE-BP in cultured cells has revealed that the IRE-BP functions either as an active aconitase, when the cells are iron-replete, or as an active RNA-binding protein, when the cells are iron-depleted. We compare properties of purified authentic cytosolic aconitase from beef liver with those of IRE-BP from tissue culture cells and establish that characteristics of the physiologically relevant form of the protein from iron-depleted cells resemble those of cytosolic aconitase apoprotein. We demonstrate that loss of the labile fourth iron atom of the Fe-S cluster results in loss of aconitase activity, but that more extensive cluster alteration is required before the IRE-BP acquires the capacity to bind RNA with the affinity seen in vivo. These results are consistent with a model in which the cubane Fe-S cluster is disassembled when intracellular iron is depleted.
铁蛋白mRNA的翻译及转铁蛋白受体mRNA的降解是由一种RNA结合蛋白，即铁反应元件结合蛋白（IRE-BP），与这些转录本中包含的被称为铁反应元件（IREs）的RNA茎环结构相互作用来调节的。在铁充足的细胞中产生的IRE-BP具有乌头酸酶（EC 4.2.1.3）活性。该蛋白与线粒体乌头酸酶具有广泛的序列同源性，并且从胞质乌头酸酶制备的肽段序列与IRE-BP的肽段相同。作为一种活性乌头酸酶，IRE-BP预计像其他乌头酸酶一样具有一个铁硫簇。这个铁硫簇被认为是该蛋白感知细胞内铁水平并相应改变IRE-BP与IREs相互作用能力的区域。在培养细胞中IRE-BP的表达表明，当细胞铁充足时，IRE-BP作为一种活性乌头酸酶发挥作用，而当细胞铁缺乏时，它作为一种活性RNA结合蛋白发挥作用。我们比较了从牛肉肝脏中纯化的天然胞质乌头酸酶与来自组织培养细胞的IRE-BP的特性，并确定来自铁缺乏细胞的该蛋白生理相关形式的特征类似于胞质乌头酸酶脱辅基蛋白的特征。我们证明铁硫簇不稳定的第四个铁原子的丢失导致乌头酸酶活性丧失，但在IRE-BP获得以体内所见亲和力结合RNA的能力之前，需要更广泛的簇改变。这些结果与一个模型一致，即在细胞内铁缺乏时，立方烷型铁硫簇会被拆解。
